There are two synapsin genes, synapsin I and II, expression of which is associated with dynamic reorganization of the neuronal cytoskeleton, axon differentiation, and synapse formation: overexpression of synapsin accelerates differentiation and depletion retards differentiation. This proposal focuses on determining the role of the actin- and microtubule- binding properties of the synapsins during neuronal differentiation. (1) To determine how depletion of one synapsin affects the distribution of the other synapsin and alters cytoskeletal dynamics, immunocytochemical and differential extraction techniques will be used to characterize the sub-cellular localization of synapsin and cytoskeletal-binding proteins in primary neuronal cultures from wildtype and synapsin knockout mice. (2) The role of the actin- and microtubule-binding domains in the trophic actions of the synapsins will be determined by (a) localizing these domains using overlay, co-sedimentation, and polymerization assays to screen mutant recombinant synapsins, (b) determining the ability of mutant synapsins to induce morphological changes in non-neuronal cells, and (c) determining if mutants can rescue morphological and developmental defects in cultures from synapsin knockout mice. (3) The role of synapsin inter/intra-molecular association in regulating actin- and microtubule-binding will be investigated. These studies will enhance our understanding of the mechanisms that control neuronal differentiation.